H.8. is a surface-exposed antigen, comprised of lipid and protein, common to Neisseria gonorrhoeae and meningitidis. H.8 antigen's restriction to the pathogenic and opportunistic Neisseria is consistent with a role in virulence. In addition, H.8 does not vary antigenically and is immunogenic in humans. This proposal is aimed at determining the structure and function of H.8 from two perspectives: i) pathogenic microbiology (as a putative virulence factor and potential immunoprophylactic); and ii) basic microbiology (as an outer membrane constituent and pathogenic variation of lipoprotein). The specific aims are i) structural characterization of H.8 antigen, using mass spectrometry and synthetic peptides; ii) elucidation the H.8 antigen, using mass spectrometry and synthetic peptides; ii) elucidation the H.8 antigen biosynthetic pathway using a) Escherichia coli clones which express and acylate H.8 antigen and b) and in vitro acylation system; and iii) functional characterization of H.8 antigen with respect to a) virulence, using genetically engineered strains in a model of systemic disease and a model of attachment and invasion; b) membrane biology, using site directed H.8 mutants to assess function in protein translocation; and c) immune responsiveness using human lymphocytes and sera with purified H.8 antigen and synthetic peptides representing continuous and discontinuous epitopes.